Adenovirus vectors have proven effective in transferring genes to the respiratory tract in vitro and in several animal species. We evaluated the efficacy of a modified type 2 adenovirus (AD2) in producing uptake and expression of the B-galactosidase reporter gene when administered by aerosol to rhesus monkeys, and evaluated the safety of this vector when linked to CFTR cDNA. Additionally we used the technique of concurrent flow spirometry to determine the dose delivered to the airway in each animal, and used a unique low pressure closed system to deliver the aerosol to spontaneously breathing animals. In two animals treated with AD2B-GAL dose dependent expression was evident throughout the lower airway. In animals treated with AD2CFTR2 by aerosol and compared with controls, no significant changes in respiratory status were evident clinically, although one animal developed radiographic and histologic evidence of a pneumonia. Animals demonstrated significant variation in the amount of aerosol material inhaled, ranging from 17% to 97%, despite use of a standard treatment protocol. This adenoviral vector appears to be effective with minimal evidence of adverse effects. Given the variation in dose delivered, dosimetry will remain a critical element in future studies of aerosol gene transfer to the respiratory tract.